Kano A, Ohama T, Abe R, Osawa S
Department of Biology School of Science, Nagoya University, Japan.
J Mol Biol. 1993 Mar 5;230(1):51-6. doi: 10.1006/jmbi.1993.1125.
We previously reported that in Micrococcus luteus, a Gram-positive eubacterium with high genomic G + C content, certain codons ending with A did not appear in coding frames, including termination sites, and tRNAs that translate these codons were not detected. These facts suggest that at least some of them are unassigned (nonsense) codons, i.e. not assigned to any amino acid or to any stop signal. We have investigated whether AGA and AUA, universal Arg and Ile codons, respectively, are really unassigned codons by using a cell-free extract prepared from M. luteus and synthetic messenger RNAs. Translation of synthetic mRNA containing in-frame AGA codons does not result in "read-through" to codons beyond the AGA codons, i.e. translation ceases at codon AGA. Essentially the same result was obtained with mRNA containing AUA in-frame. A sucrose-gradient centrifugation profile of the reaction mixture has shown that practically all of the peptides that have been synthesized are attached to 70 S ribosomes. When in-frame AGA or AUA codons are replaced by UGA codons in mRNA, no read-through occurs beyond UGA, just as in the case of AGA or AUA. However, the synthesized peptide is released from the 70 S ribosomes. These data suggest that AGA and AUA are unassigned codons and differ from UGA in that they are not used for termination.
我们之前报道过，在藤黄微球菌（一种基因组G + C含量高的革兰氏阳性真细菌）中，某些以A结尾的密码子不出现在编码框中，包括终止位点，并且未检测到翻译这些密码子的tRNA。这些事实表明，其中至少一些是未分配（无义）密码子，即未分配给任何氨基酸或任何终止信号。我们通过使用从藤黄微球菌制备的无细胞提取物和合成信使RNA，研究了分别作为通用精氨酸和异亮氨酸密码子的AGA和AUA是否真的是未分配密码子。含有框内AGA密码子的合成mRNA的翻译不会导致“通读”到AGA密码子之后的密码子，即翻译在密码子AGA处停止。含有框内AUA的mRNA也得到了基本相同的结果。反应混合物的蔗糖梯度离心图谱表明，几乎所有合成的肽都附着在70S核糖体上。当mRNA中的框内AGA或AUA密码子被UGA密码子取代时，UGA之后不会发生通读，就像AGA或AUA的情况一样。然而，合成的肽会从70S核糖体上释放出来。这些数据表明AGA和AUA是未分配密码子，并且与UGA的不同之处在于它们不用于终止。
